Recently phyllocladane diterpenoids isolated from plant Callicarpa macrophylla Vahl (Family Verbenaceae) have been found to promote plant growth and alleviate the effects of growth retardant allelochemicals. (Singh et al., U.S. Pat. No. 6,673,749 (2004)) These novel activities pave the utility of these compounds as natural plant growth promoters and for alleviation of the growth retarding effects produced by allelochemicals. The plant growth promoters hold commercial importance in intensive agriculture and have become most attractive in the agro-business of high value crops for organic cultivation. Gibberellins (specially GA3) are the most important group among the known plant growth regulators (auxins, cytokinins, abscisic acid, gibberellins, and ethylene, etc.) used for enhancing the productivity of commercial crops. However, the high cost restricts its application to high value crops only. Among, the phyllocladane diterpenoids from Callicarpa macrophylla viz. calliterpenone (16α,17 dihydroxy-3-oxo phyllocladane) and its four derivatives (calliterpenone monoacetate, calliterpenone diacetate, iso-propylidinocalliterpenone and trihydroxy calliterpenone), the most promising effects on growth promotion of roots, shoots and seed germination were demonstrated by calliterpenone (U.S. Pat. No. 6,673,749). Thus, the economical isolation of calliterpenone is needed for commercial exploitation in agro-business.
Calliterpenone and its monoacetate were first isolated by Chaterjee et al. (Tetrahedron, (28), 4319 (1972)) from the dry powdered stem and leaves of Callicarpa macrophylla. In that method, 8 kg of dried powdered leaves were extracted for 30 hours in hexane to produce a residue (1.5 g), which on column chromatography yielded 0.01% and 0.008% calliterpenone and its monoacetate, respectively. Subramaniam et al. (Phytochemistry, (13), 306-307 (1974)) isolated calliterpenone from the chloroform extract of dried leaves by column chromatography (yield not reported). Sen Gupta et al. (Journal of Indian Chemical Society, (LIII), 218-219 (1976)) isolated calliterpenone by column chromatography of neutral fractions of rectified sprit extract (yield not reported).
In all the above reports, the isolation of calliterpenone and its monoacetate have involved extraction of plant material in various solvents followed by column chromatography. Previously from this institute, Singh & Agarwal (Phytochemistry, 37(2), 587-588 (1994) and Indian J. of Chem., (33b), 1205 (1994)) isolated calliterpenone and its monoacetate by column chromatography of deposits (14 g) obtained in water extract of fresh leaves (50 kg) yielding 0.011% and 0.006% of calliterpenone and its monoacetate, respectively. The structural studies of calliterpenone and other phyllocladane diterpenoids have been done in detail by Ahamad and Zaman (Tetrahedron Lett., 2179 (1973)), Fujita et al. (Phytochemistry, (14) 2249 (1975)), Wong et al. (Acta Crystallography, Section C (47), 906 (1991)), Agrawal et al. (Indian J. Chem. (35B), 803-805 (1996)) and Gui Liu et al. (Helvetica Chemica Acta, (86) 420-437 (2003)). The yield of calliterpenone in all the reports described so far was about 0.01% on dry wt. basis (Chaterjee et al., 1972) or on fresh wt. basis (Singh et al., 1994).
For economical isolation of calliterpenone, extraction of plant material by various organic solvents produces dark viscous residues having total extractable constituents from which isolation of calliterpenone by column chromatography produced very low yield of calliterpenone. Therefore, efforts were made to isolate calliterpenone from the water extract of plant Callicarpa macrophylla using simple methods and avoiding column chromatography. Accordingly, there is a need for improved methods for isolating calliterpenone providing higher yields.